Our lack of understanding of the molecular events related to castrate-resistant (CR) growth of prostate cancer (CaP), results in the death of approximately 27,360 American men each year from this disease. The PI's long-term goal is to develop effective therapies for treating, or preferably preventing, the emergence of CR CaP. In this proposal, we will explore our recent discovery of aberrant expression of miR-125b in prostate cancer. Our hypothesis is that the expression of miR-125b in CR CaP cells is controlled by aberrantly-activated AR, that miR-125b facilitates CR growth of CaP cells by repressing the function of the p53 network, and that miR-125b has potential as a biomarker and a therapeutic target for the management of patients with CaP. Three specific Aims will be pursued to test this hypothesis. Aim 1 is to characterize the AR-mediated regulation of miR-125b in prostate cancer. We will confirm that AR signaling regulates miR-125b and determine whether overexpressed AR is aberrantly-activated in CR cells and subsequently upregulates miR-125b. These experiments will address the issue of why CR CaP cells and advanced CaP tumors express high levels of miR-125b. Aim 2 is to characterize the functions of miR-125b in prostate cancer. We will confirm that miR-125b stimulates tumorigenesis and CR growth by repressing the expression of p53, Puma, Bak1 and p14ARF, determine the role of miR-125b in anti-apoptosis and promoting cell proliferation, and evaluate the effects of miR-125b on the level and activity of AR. Data obtained from these proposed experiments will provide a mechanistic explanation of miR-125b-mediated prostatic tumorigenesis and CR growth. Aim 3 is to characterize miR-125b as a potential biomarker or therapeutic target for prostate cancer. We will detect the frequency of aberrantly-expressed miR-125b in clinical CaP samples and in sera from CaP patients and determine if miR-125b alters the response of CaP cells to therapy. These studies will facilitate the application of this miRNA as a biomarker for CaP diagnosis and prognosis and hopefully, as a target for CaP treatment. In summary, this proposal contains both technical and conceptual innovation, and has significant translational potential. Completion of these proposed studies should obtain valuable data and provide new insights into how CaP becomes castration-resistant.